Surface Plasmon Resonance Biosensors for Highly Sensitive Detection of Small Biomolecules

Small biomolecules (< 2 kDa) are crucial to a wide range of biochemical processes in the human body. Their diverse functions can include acting as hormones, neurotransmitters and pheromones. Small molecules are typically found in the blood stream and their concentrations can often be indicative of the biochemical functioning of the organism. This is particularly true of the steroids, a major class of small molecule hormone. Measurement of steroid concentrations is commonly performed to track the progression of the fertile cycle, to detect pregnancy and to help diagnose hormonal diseases (Clark et al., 1998). Immunoassay techniques dominate the market for hormonal assays. These assay kits utilise the highly selective binding of an antibody to its target antigen to detect small molecule analytes in highly complex matrices. Typically, for small molecules, the assay will consist of a plate or tube onto which the antibody is immobilised and the sample is passed over the plate surface along with an aliquot of labelled antigen. The label used is either radiochemical (radioimmunoassay (RIA)) or enzyme-based (enzyme-linked immunosorbent assay (ELISA)). The labelled antigen occupies the un-bound sites on the immunoassay plate and the signal obtained from the label by either measuring the resultant radiation, or through enzyme catalyzed colour changes, gives a standard curve from which the concentration of the free analyte can be determined. Small molecule targets are typically found in complex matrices such as blood and saliva where high molecular mass components predominate. These include proteins that can in some cases bind to the steroid hormone of interest. Their large size relative to the target antigen can also serve to sterically impede antibody binding to the steroid. Furthermore, large molecule contaminants can also bind to immunoassay surfaces causing biofouling that can further interfere with assay results. Despite their widespread application, RIA and ELISA are practically limited to use in centralised laboratory environments where careful control of the experimental conditions can be exercised. They typically involve multiple steps that require trained operators and in the case of RIA they involve the use of radioisotopic labels that require specialised handling and represent a significant health hazard. Analysis is also very labour intensive and consequently quite expensive. Samples therefore must be transported to a laboratory and